Vaccines are one of the most cost-effective measures available to the health care industry for the prevention and treatment of disease. There remains, however, an urgent need to develop safe and effective vaccines and adjuvants for a variety of diseases, including those caused by or associated with infection by pathogenic agents, cancers, genetic defects and other disorders of the immune system. Publications on vaccines, for example, Rabinovich et al., Science 265, 1401-1404 (1994), state that there is still a need for safe and heat-stable vaccines that can be administered orally and that need to be administered only a few times, preferably early in life. Also preferred are combination vaccines that can protect individuals from more than one disease, as well as vaccines that do not require an adjuvant and that can elicit cell-mediated, humoral, and mucosal immunity. To date very few, if any, vaccines meet all of these criteria.
Killed or attenuated pathogens are frequently used in conventional vaccines, and particularly in vaccines against viral infections. For example, two types of influenza vaccines are presently in use. The more conventional vaccine is an inactivated vaccine (containing killed virus) that is given by injection, typically into the arm. A second vaccine, called the nasal-spray flu vaccine (sometimes referred to as LAIV for Live Attenuated Influenza Vaccine), was approved in 2003 and contains attenuated (weakened) live viruses administered by nasal sprayer. As set forth by the World Health Organization (WHO), influenza virus types A and B are both common causes of acute respiratory illnesses. Although both virus types may cause epidemics of considerable morbidity and mortality, influenza B infections are often limited to localized outbreaks, whereas influenza A viruses are the principal cause of larger epidemics, including worldwide pandemics. The influenza virus is a member of the Orthomyxo virus family, and has a wide host range, including humans, horses, dogs, birds, and pigs. It is an enveloped, negative-sense RNA virus produced in 8 RNA segments encoding 10 viral proteins. The virus replicates in the nucleus of an infected host cell. The influenza virus is most dangerous for the young and the old, or immunocompromised individuals. The virus can be propagated to high titers in chicken eggs, which serve as the vehicle for generation of virus for the production of influenza vaccines.
Influenza A viruses undergo frequent changes in their surface antigens, whereas type B influenza viruses change less frequently. Immunity following infection by one strain may not protect fully against subsequent antigenic variants. As a consequence, new vaccines against influenza must be designed each year to match the circulating strains that are most likely to cause the next epidemic. Therefore, the WHO annually collects data based on the surveillance of the most prevalent influenza strains circulating among people and makes recommendations for the influenza vaccine composition. Currently, the vaccine includes two subtypes of influenza A virus and one influenza B virus in the vaccine. The vaccine typically protects approximately 50%-80% of healthy adults against clinical disease.
Subunit vaccines, the development of which was made possible by recombinant DNA technology, have been disappointing to date, as they exhibit only limited immunogenicity. One example is the recent clinical testing of several HIV (human immunodeficiency virus) subunit vaccines which has been stopped due not only to limited efficacy of the vaccines, but also because in some cases, immunized individuals showed accelerated disease progression when they were subsequently exposed to HIV; see, for example, Cohen, Science 264:1839 (1994); and Cohen, Science 264: 660 (1994). One disadvantage of subunit vaccines, as well as of killed virus and recombinant live virus vaccines, is that while they appear to stimulate a strong humoral immune response, they fail to elicit protective cell-mediated immunity. A major conclusion at the 1994 International AIDS Conference was that there remains a need for a cytotoxic T cell-mediated response to prevent, or reduce, HIV infectivity, which to date is lacking in vaccines in the clinic. In addition, HIV vaccines tested to date have failed to elicit immunity at the mucosal surfaces where primary HIV infection occurs.
Furthermore, the only adjuvants approved for use in the United States are the aluminum salts aluminum hydroxide and aluminum phosphate, neither of which stimulates cell-mediated immunity. In addition, aluminum salt formulations cannot be frozen or lyophilized, and such adjuvants are not effective with all antigens.
Yeast cells have been used in the production of subunit protein vaccines, including some of those tested in the aforementioned HIV vaccine trials. Yeast have also been fed to animals prior to immunization to try to prime the immune response in a non-specific manner (i.e., to stimulate phagocytosis as well as the production of complement and interferon). The results have been ambiguous, and such protocols have not generated protective cell-mediated immunity; see, for example, Fattal-German et al., Dev. Biol. Stand. 77: 115-120 (1992) and Bizzini et al., FEMS Microbiol. Immunol. 2: 155-167 (1990).
Previous studies have shown the potential for using recombinant S. cerevisiae yeast as a vaccine and immunotherapy vector. See, e.g., U.S. Pat. Nos. 5,830,463 and 7,083,787, as well as U.S. Patent Publication Nos. 2004-0156858 A1 and 2006-0110755 A1. These yeast-based immunotherapeutic products have been shown to elicit immune responses that are capable of killing target cells expressing a variety of viral and cancer antigens in vivo, in a variety of animal species, and to do so in an antigen-specific, CD8+ CTL-mediated fashion. See also Stubbs et al., Nat. Med. 7:625-629 (2001) and Lu et al., Cancer Research 64:5084-5088 (2004). More specifically, other studies have shown that Saccharomyces cerevisiae are avidly phagocytosed by and directly activate dendritic cells which then present yeast-associated proteins to CD4+ and CD8+ T cells in a highly efficient manner. See, e.g., Stubbs et al. Nature Med. 5:625-629 (2001) and U.S. Pat. No. 7,083,787.
In addition to being able to interact directly with dendritic cells, yeast have a variety of other characteristics that make them an ideal platform for immunotherapy. First, multiple antigens may be engineered for expression within a single yeast strain (see, e.g., Pichuantes et al., “Expression of heterologous gene products in yeast.” In Protein Engineering—Principles and Practice, pp. 129-162, J. L. Cleland and C. S. Craik, eds., Wiley-Liss, New York (1996). These formulations share many advantages with DNA vaccines, including ease of construction and the ability to target multiple antigens. Unlike DNA vaccines, yeast-based immunotherapeutic formulations do not require extensive purification to remove potentially toxic contaminants. The U.S. Food and Drug Administration (FDA) has designated yeast as GRAS (Generally Recognized as Safe). As such, the concern over toxicity and safety that exists with other vaccine vectors does not apply to yeast-based delivery vehicles.
Despite all the existing efforts to produce efficacious vaccines, there still remains a need for vaccine compositions that are efficient at stimulating a variety of immune responses. With respect to influenza vaccines, rates of illness among children, the elderly and certain high-risk groups is still significant, and in developing countries, vaccination may be sporadic or non-existent. In industrialized countries, production of sufficient influenza vaccine to accommodate the recipient population is hampered by production problems, high expenses and the time required to produce the vaccine using current technologies. In addition, threats of new viral strains and the possibility of future pandemics have raised interest in more effective and efficiently produced influenza vaccines. Therefore, there is a need in the art for improved vaccines that provide long-lasting and effective protection against a variety of strains of influenza, and that can be produced rapidly and safely for use in humans and other animals. However, these concerns and needs are not unique to influenza vaccines, but extend to other types of vaccines, including vaccines directed to other viruses and other infectious agents.
Indeed, many pathogens, including bacteria and parasites, infect individuals in stages, presenting a different subset of antigens for the immune system to address at each stage. In addition, many pathogens have evolved a series of strategies that allow the pathogen to “hide” from or otherwise evade the immune system. Finally, as with the viruses described above, many pathogens evolve and mutate antigens, particularly those expressed or localized on their surface, and it is also possible to be infected by multiple species or strains of pathogens at the same time, all of which complicate vaccination strategies. By way of example, infection with the parasite that causes malaria, (e.g., Plasmodium falciparum or Plasmodium vivax) initially enters the body as a sporozoite through the blood stream as a result of a bite by an infected mosquito, but then quickly infects liver cells where the sporozoite undergoes radical changes to become a merozoite. The merozoite is released from the liver cell and rapidly infects red blood cells, where the parasite multiplies, differentiates, and continues to infect other cells. Accordingly, an ideal vaccine would be able to prime the immune system to recognize and destroy all stages of the parasite, whether in the blood, in the liver, or in red blood cells. However, most vaccines are unable to prevent or eradicate all infection, but instead are focused on limiting the ability of the pathogen to cause disease or be toxic to an individual, while other stages of the life cycle and quiescent infection remain unaddressed.
Therefore, for combating pandemics of infectious disease or disease caused by other agents, it is desirable to have the ability to control or influence the type of immune response elicited, such as by preferentially eliciting a cell-mediated immune response (e.g., generation of cytotoxic T cells (CTLs)), preferentially eliciting a humoral response (e.g., an antibody response), or eliciting both types of immune responses, depending on the disease or condition being prevented or treated, and/or the immune status of an individual with respect to a particular antigen or pathogen at a given time point. In addition, it would be useful to provide compositions that can stimulate an efficacious immune response with a few administrations, and that also are effective at stimulating immune responses with exposure to low levels of antigen (dose-sparing).